The studies in this Project focus on understanding the RNA-binding proteins (RBPs) and noncoding (nc)RNAs that influence energy metabolism, since the processes that generate energy become impaired with aging. In particular, we have studied the regulation of insulin production, adipogenesis, and myogenesis by RBPs and ncRNAs. Glucose homeostasis. With rising appreciation that glucose metabolism is extensively regulated at the post-transcriptional level, we recently collaborated with the Lee laboratory (Catholic University, Seoul, Korea) in studies that found HuD to be an RNA-binding protein responsible for reducing triglyceride production in pancreatic cells through the influence of HuD as an enhancer of expression of insulin-induced gene 1 (Kim et al., Biochim Biophys Acta Gene Regulatory Mechanisms, 2016). In collaboration with the Auburger laboratory, we found that the cytoplasmic protein Ataxin-2 (ATXN2), whose deficiency leads to obesity, associates with translation initiation factors. As reported (Lastres-Becker et al., Biochimica Biophysica Acta, 2016), ATXN2 is a nutritional stress-inducible modulator of mRNA translation that affects the pre-initiation complex. Mitochondria. Over the past reporting period, we have made important progress towards understanding the role and metabolism of long noncoding (lnc)RNA in mitochondrial function. Some mitochondrial lncRNAs are encoded by nuclear DNA, but the mechanisms that mediate their transport to mitochondria are poorly characterized. One such nuclear DNA-encoded lncRNA, RMRP, was found to be the target of RBPs HuR and GRSF1, which associated with RMRP and mobilized it to mitochondria. In cultured human cells, HuR bound RMRP in the nucleus and exported it to the cytosol; subsequently, GRSF1 facilitated the function of RMRP in the mitochondria. Accordingly, silencing GRSF1 impaired the import of RMRP into mitochondria and lowered oxygen consumption rates. Our findings delineate a mechanism whereby RBPs mediate the transport of nuclear DNA-encoded lncRNAs into the mitochondria (Noh et al., Genes and Development, 2016). In addition, in collaboration with the de Cabo laboratory (Di Francesco et al. Free Radical Biology and Medicine, 2016), we have reported evidence that the NAD(P)H: quinone oxidoreductase (NQO1), besides its role in defense against reactive oxidative species, was also capable of binding RNA and forming ribonucleoprotein (RNP) complexes. One of its main targets, SERPINA1 mRNA, encodes the serine protease inhibitor -1-antitrypsin, A1AT, which is associated with disorders including obesity-related metabolic inflammation, chronic obstructive pulmonary disease (COPD), liver cirrhosis and hepatocellular carcinoma. NQO1 bound SERPINA1 mRNA but did not affect SERPINA1 mRNA levels; instead, it enhanced the translation of SERPINA1 mRNA. This novel mechanism of action of NQO1 as an RNA-binding protein may help explain its pleiotropic biological effects. Tissue aging. Although our understanding of the post-transcriptional factors that influence senescence is advancing quickly, we still know relatively little about the RBPs and ncRNAs that affect the aging process itself. During the current review period, we have collaborated with Dr. Kwons laboratory to identify microRNA miR-431 as a novel miRNA showing markedly reduced abundance in aged myoblasts. Our studies showed that elevating miR-431 improved the myogenic capacity of old myoblasts, while inhibiting endogenous miR-431 lowered myogenesis. miR-431 interacted with the 3'UTR of Smad4 mRNA, which encodes one of the downstream effectors of TGF- signaling. In agreement with the low levels of miR-431 in old myoblasts, SMAD4 levels increased in this myoblast population. In an in vivo model of muscle regeneration following cardiotoxin injury, ectopic miR-431 injection greatly improved muscle regeneration and reduced SMAD4 levels. Moreover, inhibition of miR-431 also repressed the myogenic capacity of human skeletal myoblasts. This work reveals that the age-associated miR-431 plays a key role in maintaining the myogenic ability of skeletal muscle with age (Lee et al., Genes and Development, 2015). Other energy metabolism. During this review period, we also discovered that the myogenic regulatory factor (MRF) MYF5, which functions as a transcription factor in muscle progenitor cells (satellite cells) and myocytes, displayed a novel RNA-binding function. One prominent MYF5 target was Ccnd1 mRNA, which encodes the key cell cycle regulator CCND1 (Cyclin D1). Silencing MYF5 expression in proliferating myoblasts revealed that MYF5 promoted CCND1 translation and modestly increased transcription of Ccnd1 mRNA. Overexpression of MYF5 in C2C12 cells upregulated CCND1 expression while silencing MYF5 reduced myoblast proliferation as well as differentiation of myoblasts into myotubes. Thes findings led us to propose that MYF5 enhances early myogenesis in part by coordinately elevating Ccnd1 transcription and Ccnd1 mRNA translation (Panda et al., Nucleic Acids Research 2016).